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Structural analysis of a reconstituted DNA containing three histone octamers and histone H5.

Previous work has shown that DNA and the histone proteins will combine to form structures of a complex, yet definite nature. Here, we describe three experiments aimed at a better understanding of the interactions of DNA with the histone octamer and with histone H5. First, there has been some question as to whether the methylation of DNA could influence its folding about the histone octamer. To address this point, we reconstituted the histone octamer onto a 440 base-pair DNA of defined sequence at various levels of cytosine methylation, and also onto the unmethylated DNA. The reconstituted structures were probed by digestion with two different enzymes, micrococcal nuclease and DNase I. All samples were found to contain what appear to be three histone octamers, bound in close proximity on the 440 base-pair DNA. The cutting patterns of micrococcal nuclease and DNase I remain the same in all cases, even if the DNA has been extensively methylated. The results show, therefore, that methylation has little, or no, influence on the folding of this particular DNA about the histone octamer. Second, there has been concern as to whether the base sequence of DNA could determine its folding in a long molecule containing several nucleosomes, just as it does within any single, isolated nucleosome core. In order to deal with this problem, we cut the 440 base-pair DNA into three short fragments, each of nucleosomal length; we reconstituted each separately with the histone octamer; and then we digested the reconstituted complexes with DNase I for comparison with similar data from the intact 440 base-pair molecule. The results show that the folding of this DNA is influenced strongly by its base sequence, both in the three short fragments and in the long molecule. The rotational setting of the DNA within each of the three short fragments is as predicted from a computer algorithm, which measures its homology to 177 known examples of nucleosome core DNA. The rotational setting of the DNA in the 440 base-pair molecule remains the same as in two of the three short fragments, but changes slightly in a third case, apparently because of steric requirements when the nucleosomes pack closely against one another. Finally, there has been little direct evidence of where histone H5 binds within a DNA-octamer complex.(ABSTRACT TRUNCATED AT 400 WORDS)

Base Sequence

Some physical results from an impedance camera.

We discuss some physical results from an impedance camera. We placed agar blocks of various resistivities and sizes in saline. We showed that when the agar block was small or close in resistivity to the saline solution, we were unable to reconstruct it. We attempted to reconstruct an image of a fist, but were unable to achieve this due to current flowing in the arm. We concluded that three-dimensional analysis may be needed, along with more sensitive reconstruction algorithms and measurements.

Agar

Bone mineral measurement: automated determination of midvertebral CT section.

The selection of the computed tomographic (CT) section can introduce errors in reproducibility of several percent for bone mineral measurements. An algorithm for automated determination of the midvertebral CT section from survey radiographs was developed; it is based on ridge-line tracking of the vertebral end plates. The midvertebral section is defined by all points with equal perpendicular distance to the regression lines fitted through these end-plate contours. The algorithm can be corrected by manual interaction in case of failure. It was tested on T-12 to L-4 vertebrae on 25 randomly selected localizer radiographs. Of the 250 endplate contours, 223 (89.2%) were tracked automatically; the remaining 27 (10.8%) required user interaction for successful completion.

Algorithms

The clinical algorithm nosology: a method for comparing algorithmic guidelines.

Concern regarding the cost and quality of medical care has led to a proliferation of competing clinical practice guidelines. No technique has been described for determining objectively the degree of similarity between alternative guidelines for the same clinical problem. The authors describe the development of the Clinical Algorithm Nosology (CAN), a new method to compare one form of guideline: the clinical algorithm. The CAN measures overall design complexity independent of algorithm content, qualitatively describes the clinical differences between two alternative algorithms, and then scores the degree of similarity between them. CAN algorithm design-complexity scores correlated highly with clinicians' estimates of complexity on an ordinal scale (r = 0.86). Five pairs of clinical algorithms addressing three topics (gallstone lithotripsy, thyroid nodule, and sinusitis) were selected for interrater reliability testing of the CAN clinical-similarity scoring system. Raters categorized the similarity of algorithm pathways in alternative algorithms as "identical," "similar," or "different." Interrater agreement was achieved on 85/109 scores (80%), weighted kappa statistic, k = 0.73. It is concluded that the CAN is a valid method for determining the structural complexity of clinical algorithms, and a reliable method for describing differences and scoring the similarity between algorithms for the same clinical problem. In the future, the CAN may serve to evaluate the reliability of algorithm development programs, and to support providers and purchasers in choosing among alternative clinical guidelines.

Algorithms

[Image analysis of morphologic features of the corneal endothelium including hexagonality].

We developed an analysis system for the evaluation of corneal endothelial photographs. Morphological parameters including hexagonality were analyzed semiautomatically with this system. The image analyzer used consists of an image processor MC68000 and image memories 512 x 512 x 17bytes in size. The algorithm that measures the number of apices of each cell is as follows. First, all crossing points of cell borders are detected. Then, each cell is enlarged in order by three pixels in eight directions. The number of crossing points which are overlapped with an enlarged cell is detected as the number of apices of the cell. The analysis time was 7.44 +/- 1.48 min. (mean +/- standard deviation) for 40 endothelial photographs which needed no manual trace for contrast enhancement.

Cell Count

Brainstem auditory-evoked potentials.

Brainstem auditory-evoked potentials (BAEPs) are generated in the ear and brainstem nuclei of the ascending auditory pathways following a transient acoustic stimulus. Because they can be recorded noninvasively in humans, BAEPs have a number of clinical and research applications. This paper reviews the properties of BAEPs, with particular emphasis on those characteristics that are relevant to the acquisition and analysis of the responses. Theories of the neural origins of the responses are reviewed. The dependence of the responses on the stimulus waveform and the problem of stimulus artifact are considered. Then, origins of the background noise are discussed, and the use of linear filtering and methods of artifact detection to improve the signal-to-noise ratio are reviewed. Finally, the problem of identifying parameters to quantify the responses is considered. The definition of response components in terms of response peaks and data on intra- and intersubject variability are reviewed, and the use of algorithms to measure parameters is discussed.

Acoustic Stimulation

Error measures for objective assessment of scene segmentation algorithms.

Scene segmentation is an important element in pattern recognition problems. Previous efforts to evaluate and compare scene segmentation procedures have been largely subjective. Quantitative error measures would facilitate objective comparison of scene segmentation algorithms. A theoretical discussion leading to a new generalized quantitative error measure, G2, based on comparison of both pixel class proportions and spatial distributions of "true" and test segmentations, is presented. This error measure was tested on 14 manual segmentations and 40 gynecologic cytology specimens segmented with five different scene segmentation techniques. Results indicate that G2 seems to have the desirable properties of correlation with human observation, categorization of error allowing for weighting, invariance with picture size and ease of computation necessary for a useful scene segmentation error measure.

Autoanalysis

Transducer characterization from pressure amplitude distribution measurements using a Kalman filter as parameter estimation algorithm.

The amplitude and frequency contents of a received ultrasound pulse depends on the spatial pressure amplitude distribution of the sound field, as produced by the transducer in the medium, and on the position and orientation of the reflecting surface in this field. Often, the geometry of the reflecting surface and the acoustical properties of the medium are known or can be estimated. Then it is practical to determine the transducer parameters in order to calculate the distortion of a reflected ultrasound pulse. Apart from geometrical parameters such as size and mechanical focusing, the surface velocity amplitude distribution (SVAD) of a transducer is of major importance. For some transducer configurations this SVAD may be described with a limited number of parameters. This paper presents a method, based on a Kalman filter algorithm, to assess the transducer parameters by measurement of the spatial sound field pressure amplitude distribution in various planes for different emission frequencies. Comparison of the measured pressure amplitudes with those calculated using the estimated values of the parameters shows that this method yields reliable results.

Acoustics

Fast on-line polarity correlation algorithms for muscle fibre conduction velocity measurement.

Polarity cross-correlation is a useful technique for the measurement of muscle fibre conduction velocity using surface electromyography. Owing to the nature and volume of computation involved in the correlation function, standard techniques for its estimation by a microprocessor are too slow for an on-line application. In this paper two algorithms suitable for on-line estimation of polarity function are presented. Some useful features of the correlation function, as well as careful programming and careful choice of instructions, made it possible to use a standard microprocessor to achieve higher sampling rates than those reported recently.

Action Potentials

Estimation of the post mortem period by multiple-site temperature measurements and the use of a new algorithm.

The estimation of the post mortem period, in cases where death occurred under suspicious circumstances, is usually attempted using temperature measurements taken at a single body site. Early investigations of the validity of such an approach use the abdominal skin surface, the axilla and the rectum as measurement sites (B.H. Knight, Forensic Sci. Int., 36 (1987) 47-55). However, it has recently become more common to use the rectum alone, though the ear and the nasal passages have also been utilized. Whatever site is employed, the estimates are frequently found to be inaccurate. There are several fundamental reasons for these inaccuracies, the most prominent being the unknown variation in the ambient temperature between the time of death and the commencement of measurements, and the unknown body temperature at the time of death. This paper proposes a method of overcoming the above difficulties by taking a series of measurements concurrently at a number of body sites, a technique used by several previous workers (B.H. Knight, Forensic Sci. Int., 36 (1987) 47-55). Initial investigations have shown that an improved estimation of post mortem period is obtainable by the application of a suitable decision-making algorithm.

Algorithms

[The use of the impedance measurement technic for calculating body fat mass. A validity study].

The body composition of water and fat may be calculated on the basis of measurements of its impedance. Calculations are based on algorithms developed by all measurements of the body compositions of a reference material, by means of standard methods (eg hydrodensiometry, total body water by isotope dilutimetry). A commencial impedance analyser with built-in algorithm for calculation of total body water and body fat mass was evaluated and compared with measurements of the total body water in 42 young men and women. Very good agreement was found (mean values for total water were 34.2 and 33.5, respectively, for women and 46.8 and 47.0, respectively, for men). These calculations of total body water correlated better with the reference values than calculations from four other algorithms published in the literature. Simple anthropometric measurements (skin-fold thickness, BMI) resulted in significantly higher calculations of body fat mass than calculations based on impedance measurement. This study has shown that the technique of impedance measurement has good reliable validity for calculation of the body fat mass and that, on account of its simplicity, the method is suitable for field studies and also individual measurements.

Adipose Tissue

Clinical measurement of renal clearance.

The importance of renal clearance in uronephrology has motivated the continuous reassessment of well-established methods and the relentless search for better techniques. Concerning radionuclide methods, data in the current literature confirm the validity of the simplified one-compartment model or the empirical single-sample method for renal clearance measurement, and new algorithms have been described to broaden the application of the techniques to the pediatric population. Based on the same principles, accurate measurement of renal clearance can also be obtained using contrast agents. Using the technique for determination of the glomerular filtration rate during radiographic contrast examination is recommendable. However, potential adverse effects of the contrast substances should be compared with those of other methods before using this technique for measuring renal clearance without scheduled contrast roentgenographic examination. Controversy remains concerning the validity of predicting glomerular filtration rate from plasma creatinine. Although the predicted clearance is correlated with reference clearance, analysis of individual results reveals that important differences of up to 50 mL/min frequently occur.

Algorithms

A predictive and feedback control algorithm maintains a constant glucose concentration in fed-batch fermentations.

A combined predictive and feedback control algorithm based on measurements of the concentration of glucose on-line has been developed to control fed-batch fermentations of Escherichia coli. The predictive control algorithm was based on the on-line calculation of glucose demand by the culture and plotting a linear regression to the next datum point to obtain a predicted glucose demand. This provided a predictive "coarse" control for the glucose-based nutrient feed. A direct feedback control using a proportional controller, based on glucose measurements every 2 min, fine-tuned the feed rate. These combined control schemes were used to maintain glucose concentrations in fed-batch fermentations as tight as 0.49 +/- 0.04 g/liter during growth of E. coli to high cell densities.

Acetates

Measured attenuation correction using the Maximum Likelihood algorithm.

Quantitative determination of local radioactivity concentration in positron emission tomography (PET) requires a good attenuation correction procedure to reconstruct the emission image. Using a similar Maximum Likelihood (ML) algorithm as for the reconstruction of the emission image, a method is proposed to reconstruct a transmission image, i.e. a map of absorption coefficients. This reconstructed transmission image is then used to calculate the attenuation correction factors needed for the ML reconstruction of the emission image. This approach takes automatically into account the convolution step in the acquisition process (caused by various smoothing factors, e.g. the detector width). This results in appreciable noise suppression without loss of resolution due to filtering, thus making the reconstructed images easier to interpret. A comparison is made with other estimates for the measured attenuation correction using phantom studies.

Algorithms

Ellipse test for the reduction of false positive signals in automated cytology.

One of the major problems in automated cytology is the elimination of false positive 'abnormal cell' alarms caused by objects such as overlapping cell pairs, leukocyte clusters, etc. The paper describes an algorithm for the separation of images of abnormal cell nuclei and non=nuclear objects in computer image analysis systems for automated cytology. The algorithm involves the measurement of the agreement between the object outline and a computer ellipse of equal area, aspect ratio and orientation. Results obtained with the CERVISCAN experimental computer image analysis system show that the algorithm gives good discrimination between abnormal cell nuclei and typical non-nuclear objects found in cervical scrape specimens prepared specially for automated analysis.

Cell Nucleus

Statistical evaluation of cell kinetic data from DNA flow cytometry (FCM) by the EM algorithm.

Flow cytometric DNA measurements yield the amount of DNA for each of a large number of cells. A DNA histogram normally consists of a mixture of one or more constellations of G0/G1-, S-, G2/M-phase cells, together with internal standards, debris, background noise, and one or more populations of clumped cells. We have modelled typical DNA histograms as a mixed distribution with Gaussian densities for the G0/G1 and G2/M phases, an S-phase density, assumed to be uniform between the G0/G1 and G2/M peaks, observed with a Gaussian error, and with Gaussian densities for standards of chicken and trout red blood cells. The debris is modelled as a truncated exponential distribution, and we also have included a uniform background noise distribution over the whole observation interval. We have explored a new approach for maximum-likelihood analyses of complex DNA histograms by the application of the EM algorithm. This algorithm was used for four observed DNA histograms of varying complexity. Our results show that the algorithm works very well, and it converges to reasonable values for all parameters. In simulations from the estimated models, we have investigated bias, variance, and correlations of the estimates.

Algorithms

Structural analysis of polymers of sickle cell hemoglobin. I. Sickle hemoglobin fibers.

The structure of fibers of deoxyhemoglobin S has been under investigation for several years and a number of different models have been proposed for the arrangement of molecules within the particles. We have used reconstruction and modeling techniques in our analysis of these structures. Several new approaches have been employed in this analysis in order to provide improved estimates of the co-ordinates, pairing, and polarity of the hemoglobin S molecules. Fibers have a variable pitch and, in order to minimize distortions in the reconstructed density maps associated with these variations in pitch, we have developed an iterative procedure to measure the instantaneous pitch and have modified the reconstruction algorithm to incorporate the measured values. This procedure improves the accuracy with which the hemoglobin S molecules can be located in the density maps. Furthermore, the determination of the instantaneous pitch allows us to measure directly the rotation of the individual hemoglobin molecules. These measurements are in excellent agreement with the values predicted using a random angular walk model (as originally proposed for F-actin) to describe the variable pitch. The reconstructions confirm that the fiber consists of 14 strands of hemoglobin S arranged in a hexagonally shaped cross-section. We have determined the pairing of the molecules to form double strands directly from the density maps by identifying the molecules that have intermolecular distances that conform to those of double strands in the Wishner-Love crystal. The seven double strands identified in this manner are consistent with the strand pairings proposed by Dykes et al. (1979) rather than the alternate pairings proposed by Rosen & Magdoff-Fairchild (1985). In addition, we have for the first time determined the polarity of the double strands directly from the reconstruction data. This was achieved using a procedure that amounts to essentially "dissecting" individual double strands from the reconstructed density maps so that their density distribution could be examined independently of the neighboring double strands. Knowledge of the relative polarities of the double strands is essential for determining the intermolecular interactions that stabilize the fiber.

Crystallography